Bovine pneumonic pasteurellosis, also known as shipping fever, is a major cause of sickness and death in the feedlot cattle industry (Martin, S. W. et al. Can. J. Comp. Med. 1980, 44:1-10; Yates, W. D. G. Can. J. Comp. Med. 1982, 46:225-263). The principal microorganism associated with this disease is Mannheimia (Pasteurella) haemolytica A1. It has been shown that M. haemolytica produces a heat labile cytotoxin which is specific against ruminant leukocytes (Kaehler, K. L. et al. Am. J. Vet. Res. 1980, 41:1690-1693; Shewen, P. E. et al. Infect. Immun. 1982, 35:91-94). This leukotoxin has been implicated as a major virulence factor in the pathogenesis of M. haemolytica. Its mode of action has been shown to be the impairment of the primary lung defense mechanism (inactivation of alveolar macrophages) and the induction of inflammation as a consequence of leukocyte lysis.
In the past, vaccination against M. haemolytica infection has been attempted using both live and heat-killed bacteria of various serotypes. It has been demonstrated that vaccination with heat-killed bacteria may actually enhance the development of pneumonia after challenge exposure (Sanford, S. E. Mod. Vet. Prac. 1984, 65:265-268). Immunization with live M. haemolytica vaccines have generally been unsuccessful because of low antigenicity of M. haemolytica cells and rapid inactivation by the healthy animal (Henry, C. W. Vet. Med. 1984, 1200-1206). The cytotoxic supernatant from M. haemolytica has also been used as a vaccine. This preparation contains numerous soluble antigens from the bacterium, some of which may be important in protection. Development of vaccines from the crude cytotoxic supernate requires the purification and characterization of these antigens which is difficult and costly.
Advances in molecular biology have allowed the characterization, isolation and expression of the particular genes which code for specific bacterial antigens using various recombinant DNA techniques. In fact, the gene, lktA, that codes for the full length leukotoxin (Lkt-102) of M. haemolytica has been well characterized (Lo, R. Y. C. et al. Infect. Immun., 1987, 55:1987-1996, Lo, R. Y. C. et al. U.S. Pat. No. 5,055,400). However, when the full length recombinant leukotoxin is produced in E. coli, it is very unstable and quickly degrades. The yield and recovery of the 102 kDa rLkt is therefore very poor, rendering this method of obtaining recombinant leukotoxin for use as a vaccine, costly and inefficient.
There remains a need for a highly stable derivative of the recombinant leukotoxin that retains the antigenic and immunogenic properties of the full length protein.